Andrew Roth

Behavioral and physiological effects of acute ketamine exposure in adult zebrafish

Ketamine is a non-competitive glutamatergic antagonist used to induce sedation and analgesia. In sub-anesthetic doses, it induces hyperlocomotion, impairs memory and evokes stereotypic circling in rodents. Zebrafish (Danio rerio) emerged as a promising new animal model to screen the effects of psychotropic compounds. Here, we investigated the effects of sub-anesthetic doses of ketamine on anxiety, locomotion, habituation and social behavior of adult zebrafish. Acute 20-min exposure to 20 and 40 mg/L (but not 2 mg/L) of ketamine reduced anxiety, impaired intra-session habituation, evoked circular swimming and disrupted zebrafish shoaling. Additionally, ketamine reduced whole-body cortisol levels and elevated brain c-fos expression in zebrafish. Our findings demonstrate the sensitivity of zebrafish to behavioral and physiological effects of sub-anesthetic doses of ketamine, further supporting the utility of this species as a model for neuropharmacological research, including testing ketamine and related drugs.

Automated high-throughput neurophenotyping of zebrafish social behavior.

Zebrafish (Danio rerio) are rapidly becoming an important model organism in neuroscience research, representing an excellent species to study complex social phenotypes. Zebrafish actively form shoals, which can be used to quantify their shoaling behaviors, highly sensitive to various experimental manipulations. Recent advances in video-tracking techniques have enabled simultaneous tracking of multiple subjects, previously assessed by manual scoring of animal behavior. Here we examined the effect of group-size in the shoaling paradigm (ranging from 2 to 8 fish), and evaluated the ability of novel video-tracking tools to accurately track an entire shoal, compared to traditional manual analysis of shoaling phenotypes. To further validate our approach, the effects of the psychotropic drugs lysergic acid diethylamide (LSD) and 3,4-methlenedioxymethamphetamine (MDMA), as well as exposure to alarm pheromone, previously shown to affect zebrafish shoaling, were examined. Overall, a significant difference in group size was shown in the 2-fish vs. the 3-, 4-, 5-, 6-, 7- and 8-fish groups. Moreover, both LSD and MDMA treatments reduced shoaling (assessed by increased inter-fish distance) as well as proximity (time spent together) among fish. In contrast, exposure to alarm pheromone yielded an increase in shoaling and in proximity in a time-dependent manner. Importantly, a highly significant correlation for manual vs. automated analyses was revealed across all experiments. Collectively, this study further supports the utility of zebrafish to study social behavior, also demonstrating the capacity of video-tracking technology to assess zebrafish shoaling in a high-throughput and reliable manner.

Effects of hallucinogenic agents mescaline and phencyclidine on zebrafish behavior and physiology

Mescaline and phencyclidine (PCP) are potent hallucinogenic agents affecting human and animal behavior. As their psychotropic effects remain poorly understood, further research is necessary to characterize phenotypes they evoke in various animal models. Zebrafish (Danio rerio) are rapidly emerging as a new model organism for neuroscience research. Here, we examine the effects of mescaline (5-20mg/l) and PCP (0.5-3mg/l) in several zebrafish paradigms, including the novel tank, open field and shoaling tests. Mescaline and PCP dose-dependently increased top activity in the novel tank test, also reducing immobility and disrupting the patterning of zebrafish swimming, as assessed by ethograms. PCP, but not mescaline, evoked circling behavior in the open field test. At the highest doses tested, mescaline markedly increased, while PCP did not affect, zebrafish shoaling behavior. Finally, 20mg/l mescaline did not alter, and 3mg/l PCP elevated, whole-body cortisol levels. Overall, our studies indicate high sensitivity of zebrafish models to hallucinogenic compounds with complex behavioral and physiological effects.

Perspectives of zebrafish models of epilepsy: What, how and where next?

Epilepsy is a complex brain disorder with multiple underlying causes and poorly understood pathogenetic mechanisms. Animal models have been indispensable tools in experimental epilepsy research. Zebrafish (Danio rerio) are rapidly emerging as a promising model organism to study various brain disorders. Seizure-like behavioral and neurophysiological responses can be evoked in larval and adult zebrafish by various pharmacological and genetic manipulations, collectively emphasizing the growing utility of this model for studying epilepsy. Here, we discuss recent developments in using zebrafish models to study the seizure-like behavior involved in epilepsy, outlining current challenges and strategies for further translational research in this field.

Constructing the habituome for phenotype-driven zebrafish research.

Intra-session habituation to novelty reflects spatial working memory (related to exploration and cognition), and is observed in various species, including zebrafish (Danio rerio). With the growing understanding of complex zebrafish behaviors, the extent to which they habituate remains unclear. Here we perform a large-scale characterization of zebrafish novelty-evoked (novel tank and open field) behaviors, to establish their grouping based on intra-session habituation and sensitivity to anxiolytic or anxiogenic manipulations. We also assess multiple behaviors in high- and low-anxiety sub-cohorts of a large heterogeneous zebrafish population, comparing their habituation profiles. Overall, our analyses demonstrate that anxiety responsivity and the ability to habituate show little correlation for multiple zebrafish behaviors, suggesting that they most likely represent distinct behavioral phenomena in novel environments. Using these data, we also present the habituome--a new conceptual approach to study affective and cognitive responses in zebrafish by examining a big set of their habituation phenotypes. Given marked similarity in animal novelty exploration, this approach may also be used to construct habituomes in other model organisms, including rodents and humans.

Decoding the contribution of dopaminergic genes and pathways to autism spectrum disorder (ASD).

Autism spectrum disorder (ASD) is a debilitating brain illness causing social deficits, delayed development and repetitive behaviors. ASD is a heritable neurodevelopmental disorder with poorly understood and complex etiology. The central dopaminergic system is strongly implicated in ASD pathogenesis. Genes encoding various elements of this system (including dopamine receptors, the dopamine transporter or enzymes of synthesis and catabolism) have been linked to ASD. Here, we comprehensively evaluate known molecular interactors of dopaminergic genes, and identify their potential molecular partners within up/down-steam signaling pathways associated with dopamine. These in silico analyses allowed us to construct a map of molecular pathways, regulated by dopamine and involved in ASD. Clustering these pathways reveals groups of genes associated with dopamine metabolism, encoding proteins that control dopamine neurotransmission, cytoskeletal processes, synaptic release, Ca(2+) signaling, as well as the adenosine, glutamatergic and gamma-aminobutyric systems. Overall, our analyses emphasize the important role of the dopaminergic system in ASD, and implicate several cellular signaling processes in its pathogenesis.

Alterations in grooming activity and syntax in heterozygous SERT and BDNF knockout mice: the utility of behavior-recognition tools to characterize mutant mouse phenotypes.

Serotonin transporter (SERT) and brain-derived neurotrophic factor (BDNF) are key modulators of molecular signaling, cognition and behavior. Although SERT and BDNF mutant mouse phenotypes have been extensively characterized, little is known about their self-grooming behavior. Grooming represents an important behavioral domain sensitive to environmental stimuli and is increasingly used as a model for repetitive behavioral syndromes, such as autism and attention deficit/hyperactivity disorder. The present study used heterozygous ((+/-)) SERT and BDNF male mutant mice on a C57BL/6J background and assessed their spontaneous self-grooming behavior applying both manual and automated techniques. Overall, SERT(+/-) mice displayed a general increase in grooming behavior, as indicated by more grooming bouts and more transitions between specific grooming stages. SERT(+/-) mice also aborted more grooming bouts, but showed generally unaltered activity levels in the observation chamber. In contrast, BDNF(+/-) mice displayed a global reduction in grooming activity, with fewer bouts and transitions between specific grooming stages, altered grooming syntax, as well as hypolocomotion and increased turning behavior. Finally, grooming data collected by manual and automated methods (HomeCageScan) significantly correlated in our experiments, confirming the utility of automated high-throughput quantification of grooming behaviors in various genetic mouse models with increased or decreased grooming phenotypes. Taken together, these findings indicate that mouse self-grooming behavior is a reliable behavioral biomarker of genetic deficits in SERT and BDNF pathways, and can be reliably measured using automated behavior-recognition technology.

Towards high-throughput phenotyping of complex patterned behaviors in rodents: focus on mouse self-grooming and its sequencing.

Increasingly recognized in biological psychiatry, rodent self-grooming is a complex patterned behavior with evolutionarily conserved cephalo-caudal progression. While grooming is traditionally assessed by the latency, frequency and duration, its sequencing represents another important domain sensitive to various experimental manipulations. Such behavioral complexity requires novel objective approaches to quantify rodent grooming, in addition to time-consuming and highly variable manual observation. The present study combined modern behavior-recognition video-tracking technologies (CleverSys, Inc.) with manual observation to characterize in-depth spontaneous (novelty-induced) and artificial (water-induced) self-grooming in adult male C57BL/6J mice. We specifically focused on individual episodes of grooming (paw licking, head washing, body/leg washing, and tail/genital grooming), their duration and transitions between episodes. Overall, the frequency, duration and transitions detected using the automated approach significantly correlated with manual observations (R=0.51-0.7, p<0.001-0.05). This data validates the software-based detection of grooming, also indicating that behavior-recognition tools can be applied to characterize both the amount and sequential organization (patterning) of rodent grooming. Together with further refinement and methodological advancement, this approach will foster high-throughput neurophenotyping of grooming, with multiple applications in drug screening and testing of genetically modified animals.

Experimental models for anxiolytic drug discovery in the era of omes and omics

Introduction: Animal behavioral models have become an indispensable tool for studying anxiety disorders and testing anxiety-modulating drugs. However, significant methodological and conceptual challenges affect the translational validity and accurate behavioral dissection in such models. They are also often limited to individual behavioral domains and fail to target the disorders real clinical picture (its spectrum or overlap with other disorders), which hinder screening and development of novel anxiolytic drugs. Areas covered: In this article, the authors discuss and emphasize the importance of high-throughput multi-domain neurophenotyping based on the latest developments in video-tracking and bioinformatics. Additionally, the authors also explain how bioinformatics can provide new insight into the neural substrates of brain disorders and its benefit for drug discovery. Expert opinion: The throughput and utility of animal models of anxiety and other brain disorders can be markedly increased by a number of ways: i) analyzing systems of several domains and their interplay in a wider spectrum of model species; ii) using a larger number of end points generated by video-tracking tools; iii) correlating behavioral data with genomic, proteomic and other physiologically relevant markers using online databases and iv) creating molecular network-based models of anxiety to identify new targets for drug design and discovery. Experimental models utilizing bioinformatics tools and online databases will not only improve our understanding of both gene–behavior interactions and complex trait interconnectivity but also highlight new targets for novel anxiolytic drugs.

The Zebrafish Neurophenome Database (ZND): A Dynamic Open-Access Resource for Zebrafish Neurophenotypic Data

Zebrafish (Danio rerio) are widely used in neuroscience research, where their utility as a model organism is rapidly expanding. Low cost, ease of experimental manipulations, and sufficient behavioral complexity make zebrafish a valuable tool for high-throughput studies in biomedicine. To complement the available repositories for zebrafish genetic information, there is a growing need for the collection of zebrafish neurobehavioral and neurological phenotypes. For this, we are establishing the Zebrafish Neurophenome Database (ZND; www.tulane.edu/∼znpindex/search ) as a new dynamic online open-access data repository for behavioral and related physiological data. ZND, currently focusing on adult zebrafish, combines zebrafish neurophenotypic data with a simple, easily searchable user interface, which allow scientists to view and compare results obtained by other laboratories using various treatments in different testing paradigms. As a developing community effort, ZND is expected to foster innovative research using zebrafish by federating the growing body of zebrafish neurophenotypic data.